ABSTRACT The kidney transplant waitlist comprises 83% of the overall U.S. organ transplant waitlist ? yet every year, thousands of potentially transplantable kidneys are discarded. More specifically, for every 5 waitlist patients who die or become too sick for transplantation, 3 kidneys are discarded. In addition, the quality of kidney preservation is linked to graft lifespan, kidneys having only <50% graft survival after 10 years. Experts estimate that extending kidney preservation time to 7 days or more would allow for current state-of-the-art immune tolerance induction protocols, which are showing good promise in live kidney donation at three clinical centers (MGH/Harvard, Stanford, and Northwestern), to be used in the context of deceased organ donation to reduce or eliminate the need for lifelong immunosuppression, improve the lives of kidney transplant recipients and save the healthcare system over $100 million dollars each year. Using carefully optimized protocols and effective cryoprotective agents (CPAs) developed over a decade, our team and others have successfully cryopreserved multiple tissues in an ice- free vitreous ?glassy? or ?amorphous? state, allowing for indefinite storage. However, these advances in ice-free preservation have not been matched by similar advances to prevent damage from ice growth and/or fracturing and cracking during rewarming. Recently, however, our collaborator Dr. Bischof demonstrated a scalable and biocompatible nanowarming technology using radiofrequency (RF)-excited iron oxide nanoparticles (IONPs) for ice and fracture avoidance. In this proposed study we will develop a comprehensive solution for vitrifying and nanowarming kidneys by combining Dr. Bischof's expertise with 1) our deep expertise in organ perfusion, 2) our experience with successful commercialization (Dr. Taylor, Mr. Brassil and Dr. Baicu are lead scientists behind the world market-leading organ perfusion system LifePort that enabled the IPO of Organ Recovery Systems), 3) our recent experience adapting a perfusion device for sub-zero perfusion of CPAs loaded with IONPs, and 4) our successful experiments loading rat livers with IONPs. Specifically, we will begin with precision cut rat kidney slices (PCKS), which will lend itself to high throughput assessment of CPA toxicity, CPA permeation, and critical cooling and warming rates (Specific Aims 1 and 2). These will provide the basis for studies of whole rat kidneys using CT and MRI for CPA preconditioning and IONP permeation assessment and, in our final step, vitrification and RF nanowarming with functional testing of whole rat kidneys (Specific Aims 3 and 4). This proposal will provide the basis for large animal studies in Phase 2 and subsequent extension to the cryopreservation and nanowarming of human kidneys. Altogether, our proposed research constitutes an important standalone project whose sub- strategies will likely enable further breakthroughs in biopreservation, tissue engineering, bioemedical research, and clinical practice.